Cerebral malaria is the deadliest form of malaria, potentially killing half of its victims-primarily children in sub- Saharan Africa. However, most children in malaria-endemic regions appear to develop immunity to severe malaria early in life, a phenomenon that correlates with the production of antibodies to malaria parasite antigens that are expressed on the surface of infected erythrocytes. These variant surface antigens (VSAs) are the most polymorphic families within the parasite genome: Plasmodium falciparum erythrocyte membrane proteins (PfEMP1), repetitive interspersed family proteins (RIFINs) and the sub-telomeric variable open reading frame proteins (STEVORs). Each malaria genome has hundreds of genes encoding these diverse parasite surface antigens, of which only a small subset are expressed on the surface of a given infected erythrocyte. If a subset of VSAs is specific to cerebral malaria, they would be an important target for a malaria vaccine to protect children and travelers from the deadliest consequences of malaria. Work at our field site in rural Mali suggests that malaria parasites express a stealth subgroup of these parasite surface antigens more commonly in cerebral malaria than in milder forms of malaria. Our overall goals are to determine the VSA contribution to the pathophysiology of cerebral malaria and the importance of an individual's development of antibodies to VSAs in the acquisition of immunity to cerebral malaria. This project combines novel genomic and proteomic approaches with the use of a new animal model to measure the association between particular variant surface antigens and the development of cerebral malaria and also protective natural immunity. We will determine the genetic sequences of VSAs in cerebral malaria infections and in matched uncomplicated malaria controls in a cerebral malaria case-control study and ascertain which VSAs are found on the surface on infected erythrocytes (Aim 1). Mice will be infected with these field-derived parasites and the microvascular pathophysiological effects will be identified in the brain, including associations with expressed VSAs (Aim 2). Multiple VSA fragments will be encoded onto microarray chips, which will then be exposed to patients' sera to study antibody interactions with all of these proteins at once (Aim 3). This project introduces cutting-edge research techniques-some from outside the malaria research community-including next-generation sequencing, custom microarrays, proteomics, and a promising new animal model, to advance our understanding of cerebral malaria and the development of a cerebral malaria vaccine.